In the last year, the National Institutes of Health (U.S.A.) has allocated $36 million in a 3-year program to find 100,000 human single-nucleotide polymorphisms (SNPs) (Masood, E., 1999. Nature. 398:545–546). The SNP Consortium, a group of public and private institutions, has separately committed $45 million in an effort to identify 300,000 SNPs in two years (Wellcome, 1999). Methods currently used to identify SNPs and other mutations include single-strand conformation polymorphism (SSCP; Orita, M. et al., 1989. Proc. Natl. Acad. Sci. USA. 86:2766–2770), restriction fragment length polymorphism (RFLP; Arnheim et al., 1985. Proc. Natl. Acad. Sci. USA. 82:6970–6974), amplified fragment length polymorphism (AFLP; Yunis, I. et al., 1996. Tissue Antigens. 38:78–88), micro- and mini-satellite variation (Koreth, J. et al., 1996. J. Pathol. 178:239–248), allele-specific hybridization (Shuber, A. et al., 1997. Human Mol. Genet. 6:337–347), denaturing gradient gel electrophoresis (DGGE; Guldberg, P. and Guttler, F., 1993. Nucl. Acids Res. 21:2261–2262), DNA chips with detection by fluorescence or mass spectrometry (Chee et al., 1996. Science. 274:610–614; Cargill, M. et al., 1999. Nat. Genet. 22:231–238; Hacia, J. et al., 1999. Nat. Genet. 22:164–167; Griffin, T. et al., 1999. Proc. Natl. Acad. Sci. USA. 96:6301–6306; Li et al., 1999) and direct sequencing. Another approach is direct sequencing of the genome.
The techniques listed above all require assaying each individual separately, or small pools of at most about a dozen individuals (Trulzsch, B. et al., 1999. Biotechniques. 27:266–268). Since the cost per individual is not trivial, this makes mutation discovery in large populations very expensive. However, such studies are required for determination of low-frequency point mutations with useful statistical precision (Hagmann, M., 1999. Science. 285:21–22).
A need exists for a method of identifying low-frequency inherited point mutations in large populations.